Kinetic asymmetry of renal Na+-L-lactate cotransport. Characteristic parameters and evidence for a ping pong mechanism of the trans-stimulating exchange by pyruvate.
Brush border vesicles prepared from horse renal cortex were used to study the kinetic properties of the Na+-L-lactate carrier on the outer and inner faces of the membrane. Two methods were applied for these measurements (in the absence of an electrical gradient): a direct method using influx and efflux kinetics, and an indirect method applied to trans-stimulated influx kinetics using membrane vesicles preloaded with various pyruvate concentrations (the latter enabled us to observe simultaneously the inner and outer carrier properties). Kinetic parameters obtained by the first method have shown that under sodium lactate chemical gradient, the carrier efficiency (estimated by the ratio of k = Vm/Km) is higher for the influx than efflux, a mechanism indicating a kinetic asymmetry of the transport. This difference remains at chemical equilibrium of solute concentration. The similarity of outer and inner affinity of sodium permits one to conclude that the kinetic asymmetry of the sodium lactate transport is related to the lactate-carrier interaction and not to that of the sodium-carrier. The second method using the pyruvate trans-activation effect (under sodium chemical equilibrium) has shown an affinity of lactate (Kt(out) = 1.1 mM), about 15 times higher for the carrier in the extracellular orientation than that of pyruvate for the carrier in the intracellular orientation (Kt(pyr) = 36 mM). This method has demonstrated a ping pong mechanism for the trans-activation exchange which accounts for a selective pore carrier model like a gated channel. These asymmetric properties are related to the AS glide sequential model (A and S being Na+ and lactate, respectively) proposed previously for the Na-L-lactate cotransport and to a different accessibility of the organic solute but not of the sodium on the two membrane faces.
Mengual
R
R
Laboratoire de Biochimie, Faculté de Médecine, Nice, France.
Schlageter
M H
MH
Sudaka
P
P
D000818:Animals; D001667:Binding, Competitive; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000378:metabolism*; D006736:Horses; D007668:Kidney / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D007773:Lactates / Q000378:metabolism; D019344:Lactic Acid; D008871:Microvilli / Q000378:metabolism; D027501:Monocarboxylic Acid Transporters; D011773:Pyruvates / Q000494:pharmacology*; D019289:Pyruvic Acid; D012964:Sodium / Q000378:metabolism; D027981:Symporters
D002352:Carrier Proteins; D007773:Lactates; D027501:Monocarboxylic Acid Transporters; D011773:Pyruvates; D027981:Symporters; C056939:sodium lactate cotransporter; D019344:Lactic Acid; D019289:Pyruvic Acid; D012964:Sodium